Morphological characteristics of central demyelination and remyelination: A single‐fiber study

Abstract

The characteristics of demyelination and remyelination in the central nervous system of the cat were examined using quantitative single-fiber analysis. Internodal length, fiber diameter, and nodal gap length were measured in single fibers teased from the spinal cord of normal animals and of animals with transient experimental cord compression. Demyelination was primarily paranodal, but longer extents of myelin loss occurred. New myelin sheaths were formed by oligodendrocytes and organized into segments bounded by nodes. The internodal length remained inappropriately short for fiber diameter 6 months after compression. The findings demonstrate that the spinal cord is capable of remyelinating after injury, but whether this contributes to functional recovery remains unknown. MethodSeventeen adult cats were used, including those in the electron microscopy study described in the precedingpaper [lo]. Fourteen were subjected to acute compressive lesions of the spinal cord, and 3 were studied as histological controls. Details of the method of compressing the spinal cord at the first lumbar vertebra appear elsewhere [ l o , 131. Material for histological examination was prepared in a similar way for all 17 animals, using the method of Harrison, McDonald, and Ochoa [ 141. The 14 cats with compressive lesions were killed at the following times: 2 after 2 1 hours; 1 each after 2,3,5, and 7 days; 1 after 2 weeks, 1 after 3 weeks, 2 after 4 weeks, 1 after 5 weeks, 1 after 6 weeks, 1 after 3 monrhs, and 1 after 6 months. The spinal cord was fixed in situ by retrograde aortic perfusion using 4 to 5% glutaraldehyde in 0.15 M phosphate buffer (pH 7.4) followed by 400 ml of chromebuffered osmium tetroxide [ 15, 161. Total perfusion time was approximately 1 hour. An extensive thoracolumbar laminectomy was carried out, and the fixed spinal cord from approximately the tenth thoracic to the fourth lumbar vertebra was removed in toto and cut transversely into three equal blocks approximately 1.5 cm long. The blocks were then prepared for single-fiber studies by Ohlrich's method [17, 181. Longitudinal frozen sections 60 pm in thickness were teased in 60% glycerin under a Zeiss dissecting microscope with fine mounted needles using a modification of the technique for teasing peripheral nerve fibers [ 19,201. Great care was taken to minimize stretching and damage to the fibers .Measurements and photographs were made on isolated fibers with a Leitz light microscope using a calibrated ocular graticule. Magnifications of x250 were used to measure internodal length and x400 to measure external fiber diameter and nodal gap. In determining fiber diameter, the procedure of Williams and Kashef [21] was adopted in which the mean of the diameters measured at five equidistant points in each internode was used. The measurements

Remyelination reporter reveals prolonged refinement of spontaneously regenerated myelin

“…3B). Consistent with previous EM studies (20,37), sheath length and width were positively correlated (Fig. 3E).…”

“…3C). Previous reports have relied upon thin sheaths as a hallmark of regenerated myelin (14,20,22,40), but our data indicate that it can be surprisingly similar in thickness to normal myelin. In contrast, mean mG + SC g-ratio was significantly lower (0.70 ± 0.12; P < 0.05; Fig.…”

“…Since that time, abnormally thin or short sheaths have been presumed to represent regenerated myelin. Investigators still commonly use these morphological characteristics alone to identify and study remyelination (11,19,20,22).This approach has likely limited understanding of the endogenous regeneration process. Measuring sheath length or thickness does not empirically determine whether a myelin sheath is new, degenerating, or preserved.…”

“…There are also no known molecular markers for regenerating myelin. Much of our understanding of myelin replacement is instead derived from interpretation of small-field EM or quantification of indirect histological markers (14,(18)(19)(20)(21). Groundbreaking EM studies in the 1960s and 1970s confirmed the phenomenon of central remyelination, revealing thin, loosely wrapped sheaths that resembled immature myelin (18) as well as abnormally short myelin internodes (20) a few weeks after demyelination.…”

“…Since that time, abnormally thin or short sheaths have been presumed to represent regenerated myelin. Investigators still commonly use these morphological characteristics alone to identify and study remyelination (11,19,20,22).…”